title: Characterization of oat proteins and aggregates using asymmetric flow field-flow fractionation.
abstract: The soluble proteins and protein aggregates in Belinda oats were characterized using asymmetric flow field-flow fractionation (AF4) coupled with online UV-vis spectroscopy and multiangle light-scattering detection (MALS). Fractions from the AF4 separation were collected and further characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The AF4 fractogram of the oat extracts revealed three peaks which were determined to be monomeric forms of soluble proteins, globulin aggregates, and β-glucan, respectively. The early eluting monomeric proteins ranged in molar mass (MM) between 5 and 90 kg/mol and in hydrodynamic diameter (D h) from 1.6 to 13 nm. The MM at peak maximum of the globulin aggregate peak was found to be ∼300 kg/mol and the D h was measured to be ∼20 nm. SDS-PAGE of the collected fraction across this peak revealed two bands with MM of 37 and 27 kg/mol which correspond to the α and β subunits of globulin indicating the elution of globulin aggregates. A third peak at long retention time was determined to be β-glucan through treatment of the oat extract with β-glucanase and by injection of β-glucan standards. The amount of soluble protein was measured to be 83.1 ± 2.3 wt.%, and the amount of albumin proteins was measured to be 17.6 ± 5.7 wt.% of the total protein in the oats. The results for Belinda oat extracts show that the AF4-MALS/UV platform is capable of characterizing the physicochemical properties such as MM and hydrodynamic size distribution of proteins and protein aggregates within a complicated food matrix environment and without the need to generate protein isolates.
authors.affiliation: Lund Centre for Field-Flow Fractionation, Department of Food Technology, Engineering and Nutrition, Lund University, P.O. Box 124, 221 00, Lund, Sweden. ray.runyon@food.lth.se
authors.forename: J Ray
authors.initials: JR
authors.lastname: Runyon
authors.forename: Lars
authors.initials: L
authors.lastname: Nilsson
authors.forename: Johan
authors.initials: J
authors.lastname: Alftrén
authors.forename: Björn
authors.initials: B
authors.lastname: Bergenståhl
mesh_terms: D018554:Avena / Q000737:chemistry*; D005504:Food Analysis / Q000379:methods*; D031521:Fractionation, Field Flow / Q000379:methods*; D013058:Mass Spectrometry / Q000379:methods*; D010940:Plant Proteins / Q000032:analysis*; D012639:Seeds / Q000737:chemistry*
chemical_list: D010940:Plant Proteins